Delivery of macromolecules to targeted tissues is a major problem which has prevented realization of their full potential as pharmacological agents. By means of cell culture and animal models, in conjunction with mathematical models, we are studying the transport of macromolecules such as growth factors, ribonucleases, protein toxins, albumin, streptavidin, immunotoxins, and monoclonal antibodies. The following are recent examples: (1) Transferrin receptor is overexpressed on many neoplastic cells and hence is an important cancer target. Ribonucleases have the potential to be cytotoxic agents by degrading RNA and thus preventing protein synthesis. However, the presence of cytosolic ribonuclease inhibitor limits their cytotoxicity. Examination of the crystal structure of the complex of ribonuclease and ribonuclease inhibitor led to identification of an amino acid which makes an important contact with the inhibitor but is distal from the ribonucleolytic site. By site-directed mutagenesis of ribonucleases and chemical conjugation to transferrin, we endowed two human inhibitor- sensitive ribonucleases with 5000-fold higher cytotoxicity by increasing their resistance to inhibitor and adding a targeting function. (2) At present, it is difficult to accurately measure the volume of brain tissue that is treated during high-flow microinfusion of immunotoxin (an experimental treatment for glioblastoma multiforme). We are evaluating a number of novel CT and MRI contrast agents to determine if one of them may be used as a surrogate marker for immunotoxin and thereby allow real-time monitoring of the volume of distribution of treated tissue. (3) Spatially distributed pharmacokinetic models have been developed to simulate the experimental characteristics of in vivo radiolabeling of antibodies for imaging and treatment of metastatic cancer. These protocols involve combined use of a modified antibody and a rapidly diffusible radiolabeled substance that binds to the antibody. The model includes plasma kinetics, transcapillary transport, interstitial diffusion, cellular binding and internalization, and lymphatic flow. Recent use of finite element numerical analysis has allowed the examination of more complex geometries and boundary conditions. - immunotoxin, targeted delivery, ribonuclease, transferrin, contrast agent, microinfusion, pharmacokinetic model, numerical simulation